1. Field of the Invention
The ability for a large organic compound to specifically bind to a spatial and polar structure, an epitopic site, is the basis for a broad spectrum of analytical techniques referred to as competitive protein binding assays. These techniques are predicated on labelling an analyte which provides a detectable signal. The binding of the labelled analyte to its reciprocal member of a specific binding pair allows for discrimination between the labelled analyte bound to the reciprocal member and unlabelled analyte. By allowing for a competition in an assay medium between the analyte in the sample and labelled analyte for the reciprocal binding pair member, one can then determine the amount of analyte in the medium.
A wide variety of labels have been employed, including radioactive atoms, stable free radicals, enzymes and fluorescers. The systems may be illustrated in Murphy, J. Clin. Endocr. 27,973 (1967) and U.S. Pat. Nos. 3,690,834, 3,817,837 and 3,996,345 respectively.
Despite the sensitivity and accuracy of the many systems which are presently available, it is still desirable to provide new systems which have one or more advantages over presently available systems. Advantages include enhanced sensitivity, diminished interference from materials normally present in the medium containing the analyte, ease of manipulation, simplicity of instrumentation, and the like.
2. Description of the Prior Art
Mossbach and Mattiason, Acta Chem. Scand. 24, 2,093 (1970); BBA 253, 253 (1971) teach enhancement of rates when binding two or more enzymes to supports, where the product of the first enzyme is the substrate of the second enzyme. Similar studies have been reported by Srere et al, BNAS 70, 2,534 (1973), Hervagault, et al, Eur. J. Biochem. 51, 19 (1975) and Bouin et al, BBA 438, 23 (1976). The various enzyme combinations employed include hexokinase with glucose-6-phosphate dehydrogenase, malate dehydrogenase with citrate synthetase and pyruvate dehydrogenase; xanthine oxidase with uricase and glucose oxidase with catalase. Katichalski et al, in two papers: Adv. Enzymol. 34, 444 (1971) and J. Theo. Biol., 32, 243 (1971), discuss the phenomenon of "channeling". See also, Bryce et al, Biochem. J. 153, 571 (1976).